Along with its vital role in protecting the individual against foreign invaders, the immune system may attack the individual's own tissues, thereby producing autoimmune diseases. Another undesirable activity of the immune system is the rejection of critical transplanted organs. The ability of the immune system to produce autoimmune disease or reject allografts depends on the ability of lymphocytes, particularly activated T lymphocytes, to enter the target organ or grafted tissue. Traffic to the target is by way of blood vessels and the activated T lymphocytes must be able to enter and exit through the vessel walls. Therefore, it is reasonable to suppose that the participation of T lymphocytes in autoimmune damage or graft rejection might be prevented by measures affecting their traffic.
It was discovered recently by us that T lymphocytes expressed a heparanase enzyme that specifically attacked the glycosaminoglycan moiety of the extracellular matrix secreted by endothelial cells that line blood vessels (Naparstek, Y., Cohen, I. R., Fuks, Z., and I. Vlodavsky. Activated T lymphocytes produce a matrix-degrading heparan sulphate endoglycosidase, Nature 310: 241-243 (1984)). The presence of this enzyme was associated with the ability of autoimmune T lymphocytes to penetrate blood vessel walls and to attack the brain in a model disease called experimental autoimmune encephalomyelitis (EAE).
Furthermore, it was found that the heparanase enzyme could be inhibited by heparin and by some modified heparin molecules, such as N-desulfated, N-acetylated heparin, but not by others such as totally desulfated heparin (Table 1).
In U.S. Pat. No. 4,882,318, commonly assigned to the same assignees, a method for decreasing tumor metastasis in a mammal is disclosed, comprising the administration to a mammal having a malignant tumor of about 0.05 mg/kg/day to 0.5 mg/kg/day of intact heparin or of N-desulfated, N-acetylated heparin.
Intact heparin at anticoagulant doses was used to modify the course of EAE in guinea-pigs, but was not considered a useful treatment of disease because of the danger of hemorrhage (Chelmicka-Szorc, E., Arnason, B. G. W., Arch.Neural. 27:153 (1972)). A heparin devoid of anticoagulant activity (prepared by filtration through an antithrombin column), was used to inhibit delayed type hypersensitivity reactions to foreign antigens (Schneeberger, M. S. Sr., McCluskey, R., Greene, M. I., Rosenberg, R. D., Benacerraf, B., Cell.Immunol. 82:23 (1983)). Intact heparin at doses of 17-35 mg/kg/rat was shown to inhibit the emigration of lymphocytes from the circulation (Bradfield, J. W. B., Born, G. V. R., Nature 222:1183 (1969)). Intact heparin administered with cortisone was shown to inhibit angiogenesis and cause regression of tumors (Folkman, J., Langer, R. Lindhardt, R. J., Mandelschild, C., Taylor, S., Science 221:719 (1983)).
Compositions describing heparin together with other active ingredients are described in the literature. U.S. Pat. No. 3,636,202 (Klein) discloses the sequential administration of two compositions, wherein the first contains a combination of histamine, serotonin and heparin, and where the second contains oxytetracycline, lincomycin, and nicotinic acid. None of the five ingredients used in addition to heparin are part of the present invention.
European Patent Application EP 0114589 (Folkman et al.) describes a composition for inhibition of angiogenesis in mammals in which the active agents consist essentially of 1) heparin or a heparin fragment which is a hexasaccharide or larger, and 2) cortisone or hydrocortisone or the 11-.alpha. isomer of hydrocortisone. According to the disclosure, heparin by itself or cortisone by itself are ineffective: only the combination of both gives the desired effects. Furthermore, there is no proof in the literature that there is a connection between angiogenesis and autoimmune diseases. Folkman, on page 5 of the patent application, connects angiogenesis with psoriasis and with arthritis indicating the use of doses of 25,000 units to 47,000 units per day, i.e. about 600 units per kg patient weight.times.70 kg, while according to the present invention heparin is used alone and the total dosage is, at its upper level, 500 units/day.
U.S. Pat. No. 3,033,750 (Velluz et al.) describes the preparation of N-desulfated optionally N-acetylated heparins devoid of high anti-coagulant activity.
Horvath, J. E. et al., (1975) Low Dose Heparin and Early Kidney Transplant Function, Aust. N. Z. J. Med. Vol 5, No. 6, pp. 537-539 describe the effect of subanticoagulant doses of subcutaneous heparin on early renal allograft function. The daily dosage is high (5000 U) and the conclusion of the study is that heparin in subanticoagulant doses has no effect on early graft function or graft survival and that it may be associated with increased hemorrhagic complications.
Toivonen, M. L. et al., (1982) Rat adjuvant arthritis as a model to test potential antirheumatic agents, Meth. and Find. Exptl. Clin. Pharmacol., Vol 4, No. 6, pp. 359-363, examined the effect of heparin in high dosage (1000 U/rat) in inhibition of adjuvant arthritis in rats and found that heparin enhanced the severity of the rat adjuvant arthritis.
The use of intact heparin by itself or of some chemically modified derivatives thereof in low, subanticoagulant doses, has not been disclosed in the literature for the purposes of the present invention and some of the references teach away from the invention.